Proteolysis of the peanut allergen Ara h 1 by an endogenous aspartic protease.

Abstract

The 7S and 11S globulins of peanuts are subjected to proteolysis two days after seed imbibition, with Ara h 1 and the arachin acidic chains being among the first storage proteins to be mobilized. Proteolytic activity was greatest at pH 2.6-3 and is inhibited by pepstatin A, characteristic of an aspartic protease. This activity persists in seedling cotyledons up to at least 8 days after imbibition. In vitro proteolysis of Ara h 1 at pH 2.6 by extracts of cotyledons from seedlings harvested 24 h after seed imbibition generates newly appearing bands on SDS-PAGE. Partial sequences of Ara h 1 that were obtained through LC-MS/MS analysis of in-gel trypsin digests of those bands, combined with information on fragment size, suggest that proteolysis begins in the region that links the two cupin domains to produce two 33/34 kD fragments, each one encompassing an intact cupin domain. The later appearance of two 18 and 10/11 kD fragments can be explained by proteolysis within an exposed site in the cupin domains of each of the 33/34 kD fragments. The same or similar proteolytic activity was observed in developing seeds, but Ara h 1 remains intact through seed maturation. This is partly explained by the observation that acidification of the protein storage vacuoles, demonstrated by vacuolar accumulation of acridine orange that was dissipated by a membrane-permeable base, occurs only after germination. These findings suggest a method for use of the seed aspartic protease in reducing peanut allergy due to Ara h 1.

• Authors: Karl A Wilson, Anna Tan-Wilson
• Journal: Plant physiology and biochemistry : PPB (Plant Physiol Biochem) Vol. 96 Pg. 301-10 (Nov 2015) ISSN: 1873-2690 [Electronic] France
• PMID: 26322854 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, Non-P.H.S.)
• Copyright: Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Chemical References

• Antigens, Plant
• Ara h 1 protein, Arachis hypogaea
• Glycoproteins
• Membrane Proteins
• Plant Proteins
• Aspartic Acid Proteases

Topics

• Amino Acid Sequence
• Antigens, Plant (chemistry, metabolism)
• Arachis (growth & development, immunology)
• Aspartic Acid Proteases (metabolism)
• Electrophoresis, Polyacrylamide Gel
• Glycoproteins (chemistry, metabolism)
• Hydrogen-Ion Concentration
• Membrane Proteins
• Microscopy, Confocal
• Molecular Sequence Data
• Plant Proteins (chemistry, metabolism)
• Proteolysis
• Sequence Homology, Amino Acid